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- EMDB-3641: Unraveling the self-assembly of Pseudomonas aeruginosa XcpQ secre... -

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Basic information

Entry
Database: EMDB / ID: EMD-3641
TitleUnraveling the self-assembly of Pseudomonas aeruginosa XcpQ secretin periplasmic domain provides new molecular insights into T2SS secreton architecture & dynamics
Map data
Sample
  • Complex: Dodecameric complex of the N-terminal domain of the secretin XcpQ from Pseudomonas aeruginosa
    • Protein or peptide: Secretin N terminal domain
Function / homology
Function and homology information


protein secretion by the type II secretion system / type II protein secretion system complex / protein secretion / cell outer membrane / identical protein binding
Similarity search - Function
: / GspD-like, N0 domain / Type II secretion system protein GspD / GspD/PilQ family / Bacterial type II secretion system protein D signature. / Type II secretion system protein GspD, conserved site / NolW-like / Bacterial type II/III secretion system short domain / NolW-like superfamily / Type II/III secretion system / Bacterial type II and III secretion system protein
Similarity search - Domain/homology
Type II secretion system protein D / Secretin XcpQ
Similarity search - Component
Biological speciesPseudomonas aeruginosa (bacteria)
Methodsingle particle reconstruction / negative staining / Resolution: 30.8 Å
AuthorsDouzi B / Trinh N / Ball G / Desmyter A / Michel-Souzy S / Barbier P / Kosta A / Durand E / Cambillau C / Roussel A / Voulhoux R
CitationJournal: mBio / Year: 2017
Title: Unraveling the Self-Assembly of the XcpQ Secretin Periplasmic Domain Provides New Molecular Insights into Type II Secretion System Secreton Architecture and Dynamics.
Authors: Badreddine Douzi / Nhung T T Trinh / Sandra Michel-Souzy / Aline Desmyter / Geneviève Ball / Pascale Barbier / Artemis Kosta / Eric Durand / Katrina T Forest / Christian Cambillau / Alain ...Authors: Badreddine Douzi / Nhung T T Trinh / Sandra Michel-Souzy / Aline Desmyter / Geneviève Ball / Pascale Barbier / Artemis Kosta / Eric Durand / Katrina T Forest / Christian Cambillau / Alain Roussel / Romé Voulhoux /
Abstract: The type II secretion system (T2SS) releases large folded exoproteins across the envelope of many Gram-negative pathogens. This secretion process therefore requires specific gating, interacting, and ...The type II secretion system (T2SS) releases large folded exoproteins across the envelope of many Gram-negative pathogens. This secretion process therefore requires specific gating, interacting, and dynamics properties mainly operated by a bipartite outer membrane channel called secretin. We have a good understanding of the structure-function relationship of the pore-forming C-terminal domain of secretins. In contrast, the high flexibility of their periplasmic N-terminal domain has been an obstacle in obtaining the detailed structural information required to uncover its molecular function. In , the Xcp T2SS plays an important role in bacterial virulence by its capacity to deliver a large panel of toxins and degradative enzymes into the surrounding environment. Here, we revealed that the N-terminal domain of XcpQ secretin spontaneously self-assembled into a hexamer of dimers independently of its C-terminal domain. Furthermore, and by using multidisciplinary approaches, we elucidate the structural organization of the XcpQ N domain and demonstrate that secretin flexibility at interdimer interfaces is mandatory for its function. Bacterial secretins are large homooligomeric proteins constituting the outer membrane pore-forming element of several envelope-embedded nanomachines essential in bacterial survival and pathogenicity. They comprise a well-defined membrane-embedded C-terminal domain and a modular periplasmic N-terminal domain involved in substrate recruitment and connection with inner membrane components. We are studying the XcpQ secretin of the T2SS present in the pathogenic bacterium Our data highlight the ability of the XcpQ N-terminal domain to spontaneously oligomerize into a hexamer of dimers. Further experiments revealed that this domain adopts different conformations essential for the T2SS secretion process. These findings provide new insights into the functional understanding of bacterial T2SS secretins.
History
DepositionMar 20, 2017-
Header (metadata) releaseApr 19, 2017-
Map releaseDec 20, 2017-
UpdateApr 25, 2018-
Current statusApr 25, 2018Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1.35
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 1.35
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_3641.map.gz / Format: CCP4 / Size: 2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 4.4 Å
Density
Contour LevelBy AUTHOR: 1.35 / Movie #1: 1.35
Minimum - Maximum-1.0651999 - 3.4696348
Average (Standard dev.)0.024306923 (±0.21519959)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-40-40-40
Dimensions808080
Spacing808080
CellA=B=C: 352.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z4.44.44.4
M x/y/z808080
origin x/y/z0.0000.0000.000
length x/y/z352.000352.000352.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS-40-40-40
NC/NR/NS808080
D min/max/mean-1.0653.4700.024

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Supplemental data

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Sample components

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Entire : Dodecameric complex of the N-terminal domain of the secretin XcpQ...

EntireName: Dodecameric complex of the N-terminal domain of the secretin XcpQ from Pseudomonas aeruginosa
Components
  • Complex: Dodecameric complex of the N-terminal domain of the secretin XcpQ from Pseudomonas aeruginosa
    • Protein or peptide: Secretin N terminal domain

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Supramolecule #1: Dodecameric complex of the N-terminal domain of the secretin XcpQ...

SupramoleculeName: Dodecameric complex of the N-terminal domain of the secretin XcpQ from Pseudomonas aeruginosa
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
Molecular weightTheoretical: 322 KDa

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Macromolecule #1: Secretin N terminal domain

MacromoleculeName: Secretin N terminal domain / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Pseudomonas aeruginosa (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21 (bacteria)
SequenceString: E N S G G N A F V P A G N Q Q E A H W T I N L K D A D I R E F I D Q I S E I T G E T F V V D P R V K G Q V S V V S K A Q L S L S E V Y Q L F L S V M S T H G F T V V A Q G D Q ...String:
E N S G G N A F V P A G N Q Q E A H W T I N L K D A D I R E F I D Q I S E I T G E T F V V D P R V K G Q V S V V S K A Q L S L S E V Y Q L F L S V M S T H G F T V V A Q G D Q A R I V P N A E A K T E A G G G Q S A P D R L E T R V I Q V Q Q S P V S E L I P L I R P L V P Q Y G H L A A V P S A N A L I I S D R S A N I A R I E D V I R Q L D Q K G S H D Y S V I N L R Y G W V M D A A E V L N N A M S R G Q A K G A A G A Q V I A D A R T N R L I I L G P P Q A R A K L V Q L A Q S L D T P

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Experimental details

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Structure determination

Methodnegative staining
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration1 mg/mL
BufferpH: 8
Component:
ConcentrationName
50.0 mMTris-HclTris
150.0 mMNaClSodium chloride
StainingType: NEGATIVE / Material: Uranyl Acetate
Details: The protein was coated on carbon grid for 3 min. The grid were then washed using drop method 3 times and then incubated with Uranyl Acetate 2% for 3 min.
DetailsThis sample was monodisperse.

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Electron microscopy

MicroscopeFEI TECNAI 20
Electron beamAcceleration voltage: 200 kV / Electron source: LAB6
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm
Image recordingFilm or detector model: FEI EAGLE (2k x 2k) / Average electron dose: 2.0 e/Å2

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Image processing

Particle selectionNumber selected: 1660
Details: negative monitor contrast facilitated particle manual picking (EMAN2)
CTF correctionSoftware - Name: EMAN (ver. 2.12)
Startup modelType of model: OTHER
Details: Initial model created by EMAN based on the best 2D classes
Initial angle assignmentType: NOT APPLICABLE
Final 3D classificationNumber classes: 15 / Avg.num./class: 110 / Software - Name: EMAN (ver. 2.12)
Final angle assignmentType: NOT APPLICABLE
Final reconstructionNumber classes used: 15 / Applied symmetry - Point group: C6 (6 fold cyclic) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 30.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: EMAN (ver. 2.12) / Number images used: 1660
FSC plot (resolution estimation)

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